Challenges in developing a split drive targeting dsx for the genetic control of the invasive malaria vector Anopheles stephensi

Anopheles stephensi is a competent malaria vector mainly present in southern Asia and the Arabian Peninsula. Since 2012 it has invaded several countries of eastern Africa, being an emerging risk in causing urban outbreaks, requiring urgent efforts to develop novel and more efficient strategies for targeted vector control. With the emergence of insecticide resistance, CRISPR/Cas9-based homing gene drives have been proposed as viable alternative strategies. In this study, a geographically confineable gene drive system targeting dsx was developed (dsxgRNA). The drive, in the presence of Cas9 under the control of the zpg endogenous promoter, was able to home and spread in a super-Mendelian rate comparable to the one obtained by an autonomous drive. Although inheritance rates as high as 99.8% would spread the gene drive into wild populations, observed disruption of male and female heterozygotes’ fertility due to the disruption of the dsxF and dsxM canonical splicing in the presence of the transgene would hinder this spread. These results should be considered when proposing the viability of dsx as a target gene for a population suppression gene drives in An. stephensi.